Selective estrogen receptor (ER) modulators and downregulators (SERMs and SERDs, also refereed to as anti-hormones) have saved the lives of millions of women with ER-positive breast cancer. Unfortunately, approximately one-third of these women display intrinsic resistance to these targeted therapies, while a similar fraction of women treated with anti-hormones will develop resistance over time, resulting in recurrences of often more aggressive cancers. SERMs and SERDs inhibit the classical nuclear estrogen receptor ER?, leading to the inhibition of cell proliferation and survival. In contrast, our results have shown that these same drugs activate the 7-transmembrane spanning G protein-coupled estrogen receptor GPER, resulting in the stimulation of the pro-survival PI3K/Akt axis. Furthermore, our recent studies have suggested that this may occur through phosphorylation and inactivation of the FOXO3 pro-apoptotic transcription factor. We have identified a collection of novel compounds that display strong selectivity for either ER? or GPER. By combining these pharmacological approaches with genetic approaches, we hypothesize that acquired resistance to anti-hormones involves their activation of GPER, resulting in the inactivation of FOXO3. We propose to test this hypothesis with the following specific aims: Aim 1 will test whether GPER mediates acquired breast cancer resistance to SERMs and SERDs. Aim 2 will test whether the inactivation of FOXO3 represents a critical step in enhancing tumor cell survival in the face of ER? inhibition. Aim 3 will employ a murine model of spontaneous breast cancer to test whether blocking GPER activity in combination with tamoxifen treatment, or alternatively selectively inhibiting ER? with novel drugs, prevents acquired resistance. Significance: Completion of these aims will significantly advance our knowledge of the role of the novel estrogen receptor GPER in acquired anti-hormone resistance in breast cancer. Identifying GPER as a novel mediator in acquired anti-hormone resistance in breast cancer in combination with the use novel highly selective ligands to be evaluated in this proposal could lead to significant improvements in outcome for the150,000 women diagnosed with ER-positive breast cancer each year.